The F1-ATPase of Escherichia coli has been dissociated into its 5 subunits and completely reconstituted. We are studying the binding of substrates and inhibitors to each of the native subunits. By "mixed subunit reassociation" we determine which subunit complements the defect in an ATPase mutant. Thus we can localize the mutation to a given subunit. We plan further studies on subunit interactions in free solution. We have shown that a 15-unit, amino-terminal peptide from alpha is essential for the binding of the delta subunit to F1-ATPase. We want to isolate this peptide and study its binding properties directly. We want to see if certain groups of 2 or 3 subunits derived from F1-ATPase will bind to depleted membranes. We plan to continue purification of F1-Fo in order to dissociate and reconstitute the entire complex. We have found that external ATP causes a rapid increase in passive permeability of transformed, but not of untransformed, mouse fibroblasts. We plan to test many other transformed cell lines to see if they also respond to ATP. We want to isolate mutants resistant to ATP permeabilization. Finally, we want to find out if the permeability change is due to specific phosphorylation of a membrane protein.